Thiamine pantetheine disulfide derivatives



United States Patent Ofifice Patented May 16, 1967 This invention relates to new disulfide-type thiamine derivatives.

In its broader aspect, the invention relates to thiamine wherein R is as precedingly defined, is allowed to react with another reactant (pantetheine or derivative thereof) having the formula wherein R and R are as precedingly defined, at least one of R and R being acyl, and X is an active radical which forms a disulfide structure by reaction with the pantetheine disulfides of the formula mercapto radical of the compound having the Formula N=CNHZ GHQ 0R3 0R2 Inc-o JCIIz-N SS-CI'I2CH2NHCOCHZCIIZNHCO-CHCCH2 IHI(HJH c=o In on CH3 CH2CH2OR1 (I) wherein each of R R and R represents H or acyl, i.e. (11).. For example X stands for SO M, 40 R, RCO- derived from carboxylic acid, with up to 7 car- SOR, SCN, bon atoms, at least one of R and R being acyl, and to the preparation thereof. In a more specific aspect of T 32 the invention, R, in the RCO- group, stands for H or SO CHgOH NHCOCH CH2NHCO-CHCCH2 lower alkyl (methyl, propyl, butyl, etc.), lower alkoxycarbonyl (methoxy, ethoxy, propoxy, etc), benzoyl, furoyl, furfuroyl, etc. groups with up to 7 carbon atoms. 0R3 0R Heretofore, it has been known that pantetheine, which SOCH2CH2NHOOOH2CH2NHOO (H C-AHQ, halogenorfiy is also called Lactobacillus bulgaricus factor (LBF), is a t factor to promote the growth of certain microorganisms H3O CH3 and an important substance as an intermediate in a bio- 35 being a hydrocarbon radical having up to 7 carbon atoms chemical pathway to co-enzyme A. It is also known that Such as methyl, ethyl propyl, nhenyl, tolyl em and M is some disulfide'type thiamine derivatives having disulfide an alkali metal, univalence of alkaline earth metal or linkage between the mercapto group of a thiol type Where each of and is a hydrocarbon thiamine and an alkylmercapto compound show superior radical as previously defined for or therapeuti? effects f f with h 40 More concretely, typical. pantetheine derivatives of The ob ect of this invention is to provide the new th1 Formula (111) one of the reactants, are exemplified by amine derivatives represented by general Formula I which the following; have superior therapeutic eifects relative to both thiamine and pantetheine. Another object is to provide a method Sodium l of .di l s f m for preparing the same. Further objects will be clear potassium l of ln b ps- 1f from the description mentioned below. th i Typical thiamine pantetheine disulfides of this invention Sodium salt f -fu 1 S u1f a teth in are the following Presently Preferred Compounds? triethylammonium salt of S-sulfopantetheine;

'y-benzoyl-S-cyanopantetheine;

thiamine -benzoylpantetheine disulfide; -benzoylpantetheine; thiamine a-acetyl-y-benzoylpantetheine disulfide; 5O a-propionyl- -furoyl-S-p-tolyl-sulfopantetheine; thiamine u,' -diacetylpantetheine disulfide; u-acetyl-y-benzoylpantetheine sulfoxide; etc. thiamine u,' -dipropionylpantetheine disulfide; thiamine a-formyl-v-benzoylpantetheine disulfide; Among the pantetheine derivatives above mentioned, O-acetylthiamine 'y-benzoylpantetheine disulfide; those wherein X stands for -SO M are most advantage- O-acetylthiamine 'y-acetylpantetheine disulfide;

O-acetylthiamine a,'y-diacetylpantetheine disulfide;

O-benzoylthiamine 'y-acetylpantetheine disulfide;

O-benzoylthiamine 'y-benzoylpantetheine disulfide;

O-benzoylthiamine a,y-diacetylpantetheine disulfide;

etc.

In the preparation of compounds (I) of this invention, one reactant having the formula CHL'CHHO 1 ously employed.

The pantetheine derivatives (III) wherein X is SO M are prepared by reacting pantothenoyl ethyleneimine derivatives having the formula:

wherein R and R are as precedingly defined, with a compound having the formula M S O wherein M has the significance previously set forth.

Potassium thiocyana-te can also be used instead of M 5 0 to give the corresponding pantetheine derivatives.

The reaction of the present invention is carried out in an aqueous medium, but if desired it can be carried out in an organic solvent such as methanol, ethanol, dimethyl- 3 formamide, etc. or in a mixture of water and such orga-nic solvent.

In order to carry out the reaction continuously, it is preferable to remove the disulfide compounds of this invention from the reaction system, because otherwise the reaction may reach an equilibrium with the result that continuous reaction is hindered. To withdraw the disulfide derivatives of this invention from the reaction mixture, resort can be had to the expedient of adding another solvent which can dissolve the disulfide derivatives of this invention, but which is immiscible with the initial solvent. As such solvent, chloroform, ethylacetate, benzene, etc. can advantageously be employed, when water is the initial solvent used.

It is generally known that when thiamine or its related compounds exist in a solution, equilibrium between the thiazolium type and the thio type, to which disulfide derivatives of this invention belong, is observed, and that equilibrium shifts to the thio type under alkaline conditions. Accordingly, the reaction of this invention takes place under neutral or alkaline conditions. Especially it is preferable to carry out the reaction at an alkaline pH by adding an alkaline reagent such as sodium hydroxide, potassium hydroxide, ammonia, organic bases such as pyridine, aniline, etc.

When the pantetheine derivative having the Formula (III) wherein X is H is used as one of the reactants of this invention, it is preferable that an oxidizing agent such as iodine, hydrogen peroxide, etc. be employed. Generally, the reaction proceeds smoothly at room temperature (about 20 to 30 C.). However, when necessary, it may be carried out with cooling or heating, so as to control the reaction velocity.

The disulfide type derivatives of this invention thus prepared may be isolated, if desired. Generally, the isolation is carried out by utilizing the difference in solubility in a solvent, or difference in distribution coefficent, etc., between the objective compounds and impurities. Depending on necessity, such a means as condensation of the reaction mixture, precipitation by adding any other solvent, or recrystallization, can be carried out.

The thiamine pantetheine disulfide derivatives of the present invention all have superior thiamine activities. They are free from unpleasant odor, are well absorbed from intestines, transferred to the blood and various organs at a high thiamine level, and are not decomposed by aneurinases.

Test for thiamine activity The compounds of this invention were administered for 21 days to rats which had been fed with a thiaminefree diet. As a result of this growth test, it was found that increase in the body weight of the rats thus supplied with the compounds of this invention was the same or greater than that of the rats to whom thiamine itself was administered:

limits Thiamine y-benzoflpantetheine disulfide Thiamine a-acetyl-'y-benzoylpantetheine disulfide The products of this invention all have both thiamine activities and pantetheine activities as described in detail hereinbefore, and therefore they may safely and effectively be employed in the therapy of deficiency disorders such as beriberi, neuritis, failure of growth, necrotic lesions of the adrenal cortex, porphyrin staining of hair, graying of hair, edema, etc. and further they may have an activative effect on Krebs cycle.

They may be administered preferably orally after the manner usually employed for thiamine and in comparable amounts.

The following examples set forth presently preferred illustrative, but not limitative, embodiments of the invention. In these examples, parts by weight bear the same relation to parts by volume as do grams to milliliters. Percentages are by weight.

Example 1 To a solution of 2.4 parts by weight of 'y-benzoyh pantethenyl ethyleneimide in 10 parts by volume of ethanol, 1.6 parts by weight of sodium thiosulfate dissolved in 10 parts by volume of ethanol was added. After stirring, the resultant solution was alkaline to cresol red reagent.

To the obtained solution, 3.2 parts by volume of 2-normal hydrochloric acid was added dropwise, then the solvent was removed under reduced pressure. The residue was extracted with ethanol to remove sodium chloride.

The ethanol was evaporated under reduced pressure and the residue was dissolved in water and the remaining insoluble portion was removed by extraction with ethyl acetate.

Evaporation of water from the obtained solution gave 1.9 parts by weight of sodium salt of y-benzoyl-S-sulfopantetheine.

TABLE Body Weight (g.)

Body Weight Dose rnol/ Gain (g./rat/ Test Compound day/rat After feeding 21 Days after week) Mean 1 Before with thiaminefeeding with Standard test free diet diet containing deviation test compound None 50 89 Thiamine chloride hydrochlor 0.01 54 87 95 2. 6:1:3. 8 Do 0.02 52 131 13. 8:\:1. 0 Do 0. O4 57 87 167 26. 8:l:4. 3 Thiamine-benzoyl-pantetheine disulfide 0.01 51 87 92 1.85:1.9 Do 0.02 55 87 122 11. 45:3. 1 Do 0. 04 54 87 168 26. 9:1:7. 8 Thiamine a-acetyl-v-benzoy pantetheine disulfide 0.01 49 87 111 8. 15:2. 5 Do 0. 02 51 87 129 13. 9:1:1. 7 0. 04 51 87 169 27. 3:l:3. 5

1 All the rats died after 14.9 days on the average.

To a solution of 8.1 parts by weight of thiamine chloride hydrochloride in parts by volume of water, 2.9 parts by weight of sodium hydroxide dissolved in 8 parts by volume of water was added.

After saturating the obtained solution with sodium chloride, 1 1.5 parts by weight of sodium salt of 7-17311- zoyl-S-sulfopantetheine was added to the solution with stirring to throw down resinous precipitate immediately.

After minutes stirring the precipitate was extracted with 200- parts by volume of chloroform. The extract was washed sufiiciently with water and further extracted with dilute hydrochloric acid until the extract became acidic.

The extract was alkalified by the addition of sodium bicarbonate to precipitate oily substance, which was extracted with chloroform. The chloroform layer was washed with water, dried and evaporated. 100 parts by volume of ether was added to thus obtained residue to give 7.2 parts by weight of powdery crystals of thiamine 'y-benzoylpantetheine disulfide.

Analysis.Calcd. for C3gH4oN O7S I C, H, N, 12.69. Found: C, 54.26; H, 6.63; N, 12.87.

Example 2 To a solution of 10 parts by weight of thiamine chloride hydrochloride in 15 parts by volume of water, 3.6 parts by weight of sodium hydroxide dissolved in 10 parts by volume of water was added.

16 parts by weight of sodium salt of a-acetyl-y-benzoyl- S-sulfopantetheine was added to the resultant solution with vigorous agitation to throw down resinous precipitate immediately.

The precipitate was extracted with 150 parts by volume of chloroform, and the extract was washed with water and concentrated to leave a residue.

After being washed twice with 100 parts by volume of ethyl acetate, the residue was dissolved in dilute hydrochloric acid.

The obtained solution was further extracted with 50 parts by volume of ethyl acetate to separate insolubles therefrom and was neutralized with sodium bicarbonate to obtain the crude paste-like product of thiamine a-acetyl-y-benzoylpantetheine disulfide, which was extracted again with 100 parts by volume of chloroform. The chloroform layer was washed with water, dried and evaporated. 100 parts by volume of ether was added to thus obtained residue to give 10.5 parts by weight of powdery crystals, melting at 85 C.

AnalySis.Calcd. for C I-I O N S C, 54.52; H, 6.29; N, 11.92. Found: C, 54.51; H, 6.54; N, 11.84.

Example 3 To a solution of 3.4 parts by weight of thiamine chloride hydrochloride in 5 parts by volume of water, 12 parts by volume of 10% aqueous solution of sodium hydroxide was added, and the mixture was allowed to stand for 30 minutes. After being saturated with sodium chloride, the mixture was supplied with 4.6 parts by weight of sodium S-sulfo-a,'y-di-acetylpantetheine and 30 parts by volume of ethyl acetate.

After 10 minutes stirring, the ethyl acetate layer was separated, washed with water and extracted three times with 5 parts by volume each of 2-normal hydrochloric acid.

The hydrochloric acid layer was neutralized with sodium bicarbonate and further extracted with ethyl acetate. The extract was washed with water and dried.

Evaporation of the ethyl acetate under reduced pressure and subsequent addition of ether to the residue gave pulverous thiamine agy-diacetylp-antetheine disulfide.

Example 4 To a solution of 7 parts by weight of 'y-benzoylpantetheine in 30 parts by volume of glacial acetic acid,

1.4 parts by weight of 30% aqueous hydrogen peroxide solution was added.

After cooling with ice for one hour, the resultant mixture was allowed to stand overnight at room temperature.

After the addition of water, the reaction mixture was neutralized with sodium bicarbonate to percipitate an oily substance, which was extracted with chloroform. The chloroform layer was washed with water, dried and evaporated to yield 6 parts by weight of the crude 'ybenzoylpantetheine sulfoxide.

Thus obtained sulfoxide was dissolved in 50 parts by volume of 50% aqueous ethanol, a solution of 2.8 parts by weight of thiamine chloride hydrochloride in 15 parts by volume of water was added as well as 1 part by volume of 10% aqueous solution of sodium hydroxide, and the whole mixture was then allowed to stand overnight.

After evaporation of ethanol, water was added to the residual solution. Then the mixture was extracted with chloroform and the chloroform layer was further extracted with dilute hydrochloric acid. The extract was treated in the same manner as in Example 1 to give crystals of thiamine v-benzoylpantetheine disulfide.

Example 5 To a solution of 8.5 parts by weight of 'y-benzoylpantetheine in parts by volume of methanol were added 4 parts by volume of 10% aqueous solution of sodium hydroxide and 100 parts by volume of water. To the resultant solution, the mixture of 3.4 parts by weight of thiamine chloride, 12 parts by volume of 10% aqueous solution of sodium hydroxide and 200 parts by volume of water was added with stirring. With further vigorous stirring, iodine solution consisting of 20 parts by weight of iodine, 14 parts by weight of potassium iodide and 200 parts by volume of water was dropped into the reaction mixture to the extent that the latter was continuously colored, followed by extraction with chloroform. The extract was decolorized with sodium bisulfite, washed with water, and further extracted with dilute hydrochloric acid. Then, treatment of the extract in the same manner as in Example 1 gave crystals of thiamine 'y-benzoylpantetheine disulfide.

Example 6 To a solution of 3.8 parts by weight of O-acetylthiamine hydrochloride in 5 parts by volume of water, 12 parts by volume of 10% aqueous solution of sodium hydroxide was added.

A solution of 5 parts by weight of sodium salt of S-sulfo-v-benzoylpantetheine in 5 parts by volume of water was stirred into the above-obtained solution at room temperature, and the stirring was further continued for 15 minutes to give a paste-like precipitate, which was extracted with 30 parts by volume of chloroform.

The extract was sufiiciently washed with water, dried with anhydrous sodium sulfate, and concentrated under pressure to leave a residue. 50 parts by volume of ethyl ether was added to the residue to solidify the latter, which was filtered and recrystallized from a mixture of acetone and ethyl ether to yield 3.0 parts by weight of colorless powder of O-acetylthiamine 'y-benzoylpantet-heine disulfide.

Example 7 To a solution of 4.4 parts by weight of O-benzoylthiamine hydrochloride in 5 parts by volume of water, 12 parts by volume of 10% aqueous solution of sodium hydroxide was added.

5 parts by weight of sodium salt of S-sulfo-y-benzoylpantetheine dissolved in 5 parts by volume of water was vigorously stirred into the resultant solution at room temperature to give a resinous precipitate, which was extracted with 30 parts by volume of chloroform.

The extract was washed with water, and dried with anhydrous sodium sulfate. The chloroform layer was 7 evaporated under reduced pressure. 50 parts by volume of ethyl ether was added to the residue and left to stand for a while to give crude O-benzoylthiamine -benzoylpantetheine disulfide gradually.

The crude product was dissolved with dilute hydrochloric acid and, after the elimination of insolubles, neutralized with sodium bicarbonate to give a resinous precipitate, which was extracted with chloroform and then treated in the same manner as in Example 6 to yield 2.0 parts by weight of crystals.

Having thus disclosed the invention,

1. Thiamine 'y-benzoylpantetheine disulfide.

what is claimed 5 2,752,348 6/1956 Matsukawa et al. 260256.5

OTHER REFERENCES Sa'hashi et a1. Jour. of Biochem, vol. 41, No. 4, 1954:

10 pp. 463-467. v

2. Thiamine a-acetyl-' -benzoylpantetheine disulfide.

References Cited by the Examiner UNITED STATES PATENTS ALEX MAZEL, Primary Examiner. MARY U. OBRIEN, Assistant Examiner. 

1. THIAMINE $-BENZOYLPANTETHEIN DISULFIDE. 